Whole-genome characterization and genomic stability of Mastadenovirus F40 and F41 in pediatric gastroenteritis

BACKGROUND: Human adenovirus types 40 and 41 (HAdV-40 and HAdV-41), belonging to the species Mastadenovirus faecale, are among the leading viral causes of acute gastroenteritis in children. Despite their clinical importance, whole-genome data on circulating strains remain limited. This study investigated the epidemiological and genomic characteristics of enteric Mastadenoviruses detected in children with acute gastroenteritis. METHODS: A total of 632 stool specimens collected from hospitalized children with acute gastroenteritis were screened using a multiplex gastrointestinal real-time PCR panel. Five representative Mastadenovirus-positive samples underwent shotgun metagenomic sequencing followed by genome reconstruction, phylogenetic analysis, pairwise sequence identity comparisons, genome-wide variant analysis, amino acid substitution profiling, and recombination analysis. RESULTS: Mastadenoviruses were detected in 60/632 (9.5%) children, with the highest positivity observed among those aged 1-3 years. Co-infections occurred predominantly involving bacterial pathogens. Complete or near-complete genomes were recovered for all five sequenced isolates with high sequencing depth (2,806 × -34,120 ×), genome coverage (94.8%-100% at ≥ 10 ×), and nucleotide identity (99.19%-99.78%) relative to genotype-matched reference genomes. Phylogenetic analyses identified two HAdV-F40 and three HAdV-F41 isolates. Gene-specific phylogenies of the hexon, penton base, fiber, and DNA polymerase genes consistently confirmed genotype assignment. Pairwise sequence identity analysis and recombination analysis further supported classification with no evidence of intergenotypic recombination. CONCLUSIONS: The sequenced enteric Mastadenovirus isolates exhibited high genomic conservation with limited within-genotype diversity and no detectable evidence of recent recombination. The concordance between gene-specific phylogenies, sequence identity analyses, and whole-genome comparisons supports the robustness of current molecular typing approaches and expands the limited genomic data available for circulating HAdV-F40 and HAdV-F41 strains.